Prostate cancer is the second leading cause of cancer-related deaths among men in the United States. No biomarkers exist that reliably diagnoses or predict disease outcome in prostate cancer patients. Identification of a biomarker for advanced disease (extracapsular and metastatic disease) would serve a powerful role for clinicians in excluding men from Active Surveillance Programs who should not be observed and importantly, identifying those at highest risk for aggressive prostate disease that require a switc to multimodal treatment regimens. MicroRNAs (miRNAs) are small non-coding RNAs that play essential roles in controlling growth and differentiation in human cells. MiRNAs are often misexpressed in human prostate cancers and contribute to tumor formation and metastasis. However, it is poorly understood how miRNAs function in the prostate to influence cancer progression to aggressive, lethal disease. Our application will characterize the role of miRNAs, such as the miR-888 cluster, in promoting prostate cancer progression by exploring their use as fluid-based biomarkers and therapeutic targets for advanced forms of extracapsular prostate cancer. SPECIFIC AIMS: We will employ an innovative biomarker source for miRNA detection - expressed prostatic secretions in urine (EPS urine) - prostatic fluids that are easily obtained non-invasively in the clinic during a standard urological exam. We hypothesize that miRNAs identified in clinically annotated EPS urine specimens will directly correlate with the underlying molecular progression of prostate cancer. Indeed, our preliminary data shows that miR- 888 is elevated in EPS urine from patients with high-grade prostate cancer and can function to induce proliferation and motility of human prostate cancer cells in vitro. In this application, we will 1) validate the expression of prioritized prostate cancer-associated miRNAs, namely members of the miR-888 cluster, within a large cohort of EPS urine specimens (non-cancer, organ-confined, non-organ confined & metastatic disease) via quantitative real-time PCR analysis and determine if they can discriminate for advanced and lethal forms of prostate cancer; and 2) characterize the functional role of EPS urine- derived miRNAs, such as miR-888, during prostate cancer progression using in vitro assays and mouse models. We are in a unique position to conduct these studies since our translational group has access to one of the largest EPS urine repositories in the nation. Investigating the role of EPS urine-derived miRNAs during prostate cancer progression promises to reveal novel strategies to detect and treat this deadly disease.